To permit field-independent manipulations to be made in the ray path, it is necessary to arrange in the ray path manipulation elements designed for this purpose—e.g., phase plates—at a plane conjugate with the objective pupil. As the back focal lengths of the exit pupils of most objectives are negative, and therefore the exit pupils of the objectives are virtually inside the objective, the objective pupil in conventional one- or two-stage microscopical imaging systems for widefield microscopical imaging of a sample are inaccessible unless a relay optical system is provided.
A microscopical imaging system with a relay optical system permitting targeted pupil manipulation outside the objective is described in U.S. Pat. No. 5,808,791, for example. The relay optical system disclosed there has an objective-side and an image-side component, with a pupil modulator—e.g., a phase plate or a contrast modulation element—being arranged between these components, at a plane on which the objective pupil is imaged. The image-side component may consist of two lens units. The pupil modulation element is a static element.
U.S. Pat. No. 5,959,772 described a relay optical system with variable magnification, i.e. with a zoom function. This system transfers an intermediate image to an image plane, in which a camera may be positioned. The relay optical system consists of five lens groups. A first lens group is focused on the intermediate image and thus collimates the rays coming from there. Three other lens groups together constitute the variable power system, with the two outer groups being displaceable along the optical axis. A fifth lens group finally projects the collimated ray path onto the image plane. Pupil manipulation is not intended in this system.
U.S. Pat. No. 7,952,800 also describes a relay optical system with variable magnification. The relay system disclosed therein transfers a magnified intermediate image to an image plane; at the same time, the system is designed in such a way that zooming changes the positions of the entrance and exit pupils but minimally. The system consists of five lens groups, four of which are arranged on the objective side, i.e. between the objective and a pupil plane within the relay system, and three of these four lens groups can be displaced relative to the others and to each other to ensure the zoom function. A fifth lens group, consisting of a cemented component, is arranged on the image side, i.e. between the pupil plane and the image plane or exit pupil, respectively.
Commonly, such relay systems can only be applied with image fields smaller than those used in widefield microscopy; moreover, they are designed for a narrow spectral range only.
In recent years, furthermore, adaptive elements such as, e.g., membrane mirrors or spatial light modulators (SLMs) have continuously been improved technically, so that now they are generally applicable in commercial widefield microscopy and, in fact, are used for correcting optical aberrations, implementing special contrasting methods and fast focusing, to name but a few examples. One example of this is described in U.S. Pat. No. 7,764,433. To ensure a field-independent effect of the adaptive optical element, it is placed at a pupil plane and can be used, e.g., to correct spherical aberrations as a function of the depth of focus. For this purpose, the adaptive optical element is positioned at a plane that is conjugate with the objective plane, where the position of each light ray depends on the angle of emergence from the sample only. For this purpose, the arrangement described in U.S. Pat. No. 7,764,433 is also provided with a relay optical system, which consists of a single lens each on the objective side and on the image side, between which the adaptive optical element is arranged. In the simplest case, this is a so-called 4f system, with the relay lengths corresponding to twice the two focal lengths of the lenses of the relay optical system.
With such a system, it is not possible, though, to effect comprehensive color correction for large spectral ranges; moreover, chromatic aberrations with regard to the image field cannot be corrected satisfactorily, if at all. Moreover, the system is not designed for the use of reflective adaptive optical elements either, in which an oblique incidence to or exit from the adaptive element may cause further aberrations. Even though an adaptive optical element is employed, color-dependent aberrations will occur if the pupil position for different light colors is attained only with a finite accuracy or if the pupil size is not filled exactly; these aberrations increase in proportion with the size of the field angle, which in widefield microscopy is particularly large.